Quantitation of Gluconeogenesis by 2H Nuclear Magnetic Resonance Analysis of Plasma Glucose Following Ingestion of 2H2O

Abstract

We present a simple 2H NMR assay of the fractional contribution of gluconeogenesis to hepatic glucose output following ingestion of 2H2O. The assay is based on the measurement of relative deuterium enrichment in hydrogens 2 and 3 of plasma glucose. Plasma glucose was enzymatically converted to gluconate, which displays fully resolved deuterium 2 and 3 resonances in its 2H NMR spectrum at 14.1 T. The signal intensity of deuterium 3 relative to deuterium 2 in the gluconate derivative as quantitated by 2H NMR was shown to provide a precise and accurate measurement of glucose enrichment in hydrogen 3 relative to hydrogen 2. This measurement was used to estimate the fractional contribution of gluconeogenesis to hepatic glucose output for two groups of rats; one group was fasted for 7 h and the other was fasted for 29 h. Rats were administered 2H2O to enrich total body water to 5% over the last 4-5 h of each fasting period. For the 7-h fasted group, the hydrogen 3/hydrogen 2 enrichment ratio of plasma glucose was 0.32 ± 0.09 (n = 7). This indicates that gluconeogenesis contributed 32 ± 9% of total hepatic glucose output with glycogenolysis contributing the remainder. For the 29-h fasted group, the hydrogen 3/hydrogen 2 enrichment ratio of plasma glucose was 0.81 ± 0.10 (n = 6), indicating that gluconeogenesis supplied the bulk of hepatic glucose output (81 ± 10%).